The cytosolic glutathione S-transferase (GST) of the rabbit lung is composed primarily of two equally abundant isozymes. These isozymes, GST 25kd and GST 24kd, are homodimers that are distinguishable by 1) the mobilities of their subunits 25 and 27k daltons, respectively, in SDS polyacrylamide electrophoretic gels; 2) their isoelectric points, pI 7.4 and 9.1, respectively, by 2- dimensional gel electrophoresis; 3) by distinct immunoreactivity with polyclonal antisera elicited against each isozyme and; 4) differences in the N-terminus (GST 27kd appears to have a blocked N-terminus similar to the alpha class of GST, and GST 25kd has 15 of 17 residues in common with the pi class isozymes of other species). The specific activity of GST 27kd is half that of GST 25kd with chloro-2,4-dinitrobenzene as substrate but is twice that of GST 25kd with pyrene 4,5-oxide (PO) as substrate. The activity of both isozymes toward PO is almost non- stereoselective in contrast to the stereoselectivity of the cytosol for the S-configured prochiral carbon. The total PO activity of the purified isozymes represents at best 10% of the cytosolic activity. The subunits of GST 27kd and GST 25kd are present in heptic cytosol, which is stereospecific for the S-configured prochiral carbon of PO, but were not purified as homogeneous isozymes by the methods used for the pulmonary cytosol. However, similar to the pulmonary isozymes, the stereospecificity and the specific activity towards PO was not maintained during purification of the hepatic GST. The specific activity toward PO of the rabbit hepatic and pulmonary preparations were comparable to those of hepatic GST preparations from other species, except for the rat and human GST mu, which were 10 times more active. Whether there is an endogenous cytosolic factor that modulates the PO activity of some GST isozymes remains to be determined, in addition to studying the cellular distribution and genetic regulation of GST and the relationship, if any, to the susceptibility of the lung to toxicants/carcinogens.